Effect of bacterial satellites on Chlamydomonas reinhardtii growth in an algo-bacterial community

The growth characteristics of an algo-bacterial community (Chlamydomonas reinhardtii and bacterial satellites) were studied, as well as the mechanism and patterns of bacterial effect on algae. Four strains of predominant bacteria were isolated and partially characterized. They were assigned to the following taxa: Rhodococcus terrea, Micrococcus roseus, and Bacillus spp. A pure culture of the alga under study was obtained by plating serial dilutions on agarized media. Within the algo-bacterial association, the alga had a higher growth rate (0.76 day^?1) and yield (60 μg chlorophyll/ml culture) than in pure cultures (0.4 day^?1 and 10 μg chlorophyll/ml culture, respectively). The viability of the algal cells within the association was retained longer than in pure culture. Among the isolated bacterial satellites, strains B1 and Y1, assigned to the species Rhodococcus terrae, had the highest stimulatory effect on algal growth. The culture liquid of bacteria incubated under the conditions not permitting growth stimulated algal growth; the culture liquid of actively growing bacteria had an opposite effect.     

Effects of cytochalasin B on division and calcium carbonate extrusion in a calcareous alga.

Cytochalasin B (CB) (100 μg/ml) reversibly blocked cell division and cuased the formation of abnormal cytoplasmic bodies in the alga Cricosphaera carterae. Concentrations of 20 μg/ml and 40 μg/ml CB were without effects. In the presence of CB, calcified bodies (coccoliths) which form in Golgi vesicles and are normally extruded through the plasma membrane were not extruded and accumulated within the cell. CB appeared to alter the membranes of Golgi vesicles containing coccoliths. DMSO (10% $\text{v}\text{v}$), the solvent for CB, was without effect on cell division and coccolith extrusion. A concentration of 20% $\text{v}\text{v}$ DMSO inhibited cell division irreversibly.     

Seasonal variation in antifouling activity of crude extracts of the brown alga Bifurcaria bifurcata (Cystoseiraceae) against cyprids of Balanus amphitrite and the marine bacteria Cobetia marina and Pseudoalteromonas haloplanktis

Previous studies have demonstrated that macroalgae from Brittany (France) contain products with antifouling activity against marine bacteria, fungi, diatoms, seaweeds and mussels. Little is known regarding the ecological function of these compounds and insufficient attention has been paid to evaluating the possible temporal variation in antifouling activity. Studies of chemical defenses in both terrestrial and marine organisms suggest that organisms vary widely in the production of chemical defenses associated with physical (temperature, light) and biological (e.g. grazing pressure) factors, season and geographical location. The present study aimed to investigate the antifouling activity of crude extracts of monthly collections of the brown alga, Bifurcaria bifurcata, against two marine bacteria, Cobetia marina and Pseudoalteromonas haloplanktis, and cypris larvae of the barnacle, Balanus amphitrite. The toxicity of the extracts was determined with a B. amphitrite nauplius assay.The antimicrobial activity of the extracts was found to be subject to seasonal variation, with the highest level of activity recorded from samples collected between April and September. Results of the anti-settlement experiments showed that the extracts of B. bifurcata (when tested from 0 to 100 μg/ml) can be divided into three groups on the basis of their minimum inhibitory concentrations (MICs): (1) extracts from plants collected from September to March reduced settlement at nontoxic concentrations (50-100 μg/ml); (2) extracts from plants collected from April to July (which were the most active extracts) reduced settlement significantly when tested at >5 μg/ml, but were toxic at 100 μg/ml; (3) the extract prepared from plants harvested in August was inhibitory at >25 μg/ml, but was toxic at 100 μg/ml. Toxicity tests on nauplii showed that LC₅₀ values of samples from the September to March collections were >100 μg/ml, demonstrating that they were nontoxic to nauplii. In contrast, samples obtained from the April to August collections were toxic to nauplii; the most toxic ones being from algae collected in May (LC₅₀=55.6 μg/ml) and in June (LC₅₀=38.3 μg/ml).The antifouling activity of extracts thus reached a peak in summer corresponding to maximal values for water temperature, light intensity and fouling pressure. It remains to be investigated whether this activity has an ecological role in the alga.     

Action of two juvenile hormone antagonists on lepidopteran epidermis

The juvenile hormone antagonist ETB (ethyl-4-2(t-butylcarbonyloxy)-butoxybenzoate) caused formation of precocious larval-pupal intermediates after the 4th (penultimate)-larval instar of the tobacco hornworm, Manduca sexta, when 50 μg were applied to any 3rd stage larvae or to 4th stage larvae within 12 hr after ecdysis. This dose was most effective within 12 hr after ecdysis to the 3rd stage. In the black mutant larval assay for juvenile hormone, ETB had activity, 0.75 μg per larva giving half-maximal score. In vitro ETB acted as a juvenile hormone to prevent the ecdysteroid-induced change in commitment at concentrations above 0.1 μg/ml with an ED₅₀ at 2.8 μg/ml and as a partial juvenile hormone antagonist to 0.1 μg/ml juvenile hormone I at concentrations between 10^?3 and 10^?2 μg/ml. By contrast, EMD (ethyl-E-3-methyl-2-dodecenoate) had little juvenile hormone-like activity in vitro up to its limits of solubility (100 μg/ml) and exhibited sporadic partial juvenile hormone antagonistic activity in vitro at concentrations between 1 and 100 μg/ml. Since these concentrations were 10–1000 times that of juvenile hormone I in the medium, EMD apparently is not an efficient competitor.     

Inhibitory effect of fucoidan from brown alga Fucus evanescens on the spread of infection induced by tobacco mosaic virus in tobacco leaves of two cultivars

The effect of fucoidan from the brown alga Fucus evanescens on the spread of infection induced by tobacco mosaic virus (TMV) was investigated in the leaves of tobacco (Nicotiana tabacum L.) of two cultivars (Ksanti-nk and Samsun). In the leaves of cv. Ksanti-nk inoculated with a mixture of TMV preparation (2 μg/ml) and fucoidan (1 mg/ml), the number of local necrotic lesions induced by the virus decreased by more than 90% as compared with the leaves inoculated with the virus alone. In tobacco leaves of cv. Samsun, virulence and the concentration of the virus 3 days after inoculation with the same mixture of TMV and fucoidan were by 62 and 66%, respectively, lower than in the leaves inoculated with TMV alone. As the infection spread, the inhibitory effect of fucoidan decreased. When the leaves were treated with fucoidan before and after the inoculation with TMV, its antiviral activity was less pronounced than when a mixture of the virus and the polysaccharide was used as inoculum. Electron microscopic investigation of TMV mixed with fucoidan often showed agglutinated virions. The highest virulence of the mixture (TMV preparation, 12 μg/ml, plus fucoidan, 1 mg/ml) was observed upon its twofold dilution, and after that it decreased. It was concluded that, when the leaves were inoculated with the mixture of TMV and fucoidan, the latter affected not only the plant but the virus as well. Treatment of tobacco leaves, cv. Ksanti-nk, with actinomycin D (10 μg/ml) 24 h before the inoculation with TMV almost completely suppressed the effect of fucoidan, indicating that fucoidan acted at a gene level.     

Action of 2,4-Dichlorophenoxyacetic Acid onNostoc linckia: Impact of Glucose and Tryptophan

2,4-Dichlorophenoxyacetic acid (2,4-D) stimulated growth and heterocyst differen- tiation ofNostoc linckia in nitrogen-free medium at lower concentrations (100 μ.g/mL) while its higher concentrations inhibited both processes and 1500 μg/mL proved to be lethal. Dry mass and specific growth rate of the alga declined with increasing concentration of 2,4-D in the range of 100–1500 μg/mL. Glucose slightly increased the heterocyst frequency without any lag in their differentiation. Tryptophan promoted growth of the alga, and formation of heterocysts (nearly three-fold). Tryptophan (50 μg/mL) complex medium with 1 mg 2,4-D per mL did not produce mature heterocysts. The filaments were fragmented at the point of hererocyst development and detached heterocysts germinatedin situ. Glucose and tryptophan protected the alga, its growth and heterocyst differentiation even at the lethal concentration of the herbicide. We are grateful to the Head, Department of Botany,Banaras Hindu University, Varanasi, for providing the necessary facilities. The first author is also grateful to the Principal,K.D. College, Kutir-Chakkey, Jaunpur, for his interest in this study.     

Flame atomic emission spectrometric determination of aluminium in a bovine blood plasma matrix

A flame atomic emission spectrometric method, is described for the determination of aluminium in bovine blood plasma matrices. Plasma samples are wet-digested and solutions are aspirated into a conventional nitrous oxide-acetylene flame. Analyte emission is monitored at 396.15 nm with corrections for background emission being obtained from measurements several tenths nm on both sides of the aluminium line. The mean recovery of 0.3–5 μg/ml aluminium added to model solutions containing 500–5000 μg Na/ml, 50–1000 μg Ca/ml, 2000–5000 μg K/ml, or simulated plasma digests containing Na, K, and Ca was 100,6% (SD = 10.9, df = 60); the mean recovery of 0.3, 0.5, and 1.0 μg/ml aluminium added to blood plasma before digestion was 94.3% (SD = 9.8, df = 33) indicating no serious interferences. For standard solutions, the detection limit (signal: peak-to-peak noise = 1) was 0.02 μg/ml by flame emission, and 0.12 μg/ml by atomic absorption measurements with the same instrument. A sample taken through the analytical procedure, gave a detection limit of 0.05 μg/ml suggesting the submicrogram per milliliter region as the lower practical limit of the method.     

Effect of bovine pituitary hormone preparations on newt lens regeneration in vitro: stimulation by thyrotropin

The anterior lobe of the pituitary gland can stimulate lens regeneration from the dorsal iris in the newt Notophthalmus viridescens. We have studied the effect of pituitary hormone preparations on this process. Dorsal irises were cultured for 20 days in diluted Medium 199 supplemented with 10% fetal calf serum. Bovine thyrotropin TSH-B8 at concentrations of 30 to 3000 μg/ml significantly stimulated lens regeneration in these dorsal irises. Well-developed lenses, up to stage 9, were formed, in which γ-crystallin, a protein specific for lens fibers of young lenses, was detected by immunofluorescence. Additionally, the mitotic index was 5.5 times elevated in these explants when compared to their controls. Lutropin LH-B10 at concentrations of 30 to 3000 μg/ml, prolactin PRL-B4 at concentrations of 23 to 1600 μg/ml, and porcine adrenocorticotropin ACTH-6002 at concentrations of 3 to 300 μg/ml did not stimulate lens regeneration. A weak stimulation of lens formation was observed in iris cultures with 2700 μg/ml of follitropin FSH-B1 or 3000 μg/ml somatotropin GH-B18, but not at concentrations of 30 μg/ml. Our results suggest that the inherent ability of the dorsal iris to form lens can be activated by the bovine thyrotropin preparation TSH-B8.     

Antifungal chitinase against human pathogenic yeasts from Coprinellus congregatus

The inky cap, Coprinellus congregatus, produces mushrooms which become autolyzed rapidly to generate black liquid droplets, in which no cell wall is detected by microscopy. A chitinase (Chi2) which is synthesized during the autolytic phase of C. congregatus inhibits the growths of Candida albicans and Cryptococcus neoformans up to 10% at the concentration of 10 μg/ml, about 50% at concentration of 20 μg/ml, and up to 95% at the concentration of 70 μg/ml. Upon treatment these yeast cells are observed to be severely deformed, with the formation of large holes in the cell wall. The two yeast species show no growth inhibition at the concentration of 5 μg/ml, which means the minimum inhibitory concentrations for both yeast species are 10 μg/ml under these experimental conditions.     

Eimeria tenella: Effect of narasin,a polyether antibiotic on the ultrastructure of intracellular sporozoites

Eimeria tenella sporozoites were inoculated into cultures of chick kidney cells in the presence of 0.01 or 0.1 μg/ml of narasin and incubated at either 40 or 30 C for 24 hr. Electron microscopic examination revealed that either concentration of this polyether ionophore caused extensive ultrastructural damage to the intracellular sporozoite at 40 but not at 30 C, indicating that the severity of the coccidiocidal effect is influenced by temperature. The effect of 0.01 μg/ml monensin on the intracellular parasite was similar to that of narasin, suggesting a common destructive mechanism. The host cells were unaffected by 0.01 μg/ml of narasin at either temperature and by 0.1 μg/ml at 30 C, indicating that the polyether ionophores can be selectively lethal for the parasite. However, when the host cells were treated with 0.1 μg/ml narasin and incubated at 40 C, ultrastructural abnormalities were evident. The results suggest that the coccidiocidal effect of the polyether ionophorous antibiotics may be a general osmotic phenomenon.     

Anaerobic cellulolytic rumen fungal populations in goats fed with and without Leucaena leucocephala hybrid, as determined by real-time PCR

The effect of Leucaena leucocephala hybrid-Bahru (LLB), which contains a high concentration of condensed tannins, on cellulolytic rumen fungal population in goats was investigated using real-time PCR. The fungal population in goats fed LLB was inhibited during the first 10 days of feeding, but after 15 days of feeding, there was a tremendous increase of fungal population (157.0 μg/ml), which was about fourfold more than that in control goats (39.7 μg/ml). However, after this period, the fungal population decreased continuously, and at 30 days of feeding, the fungal population (50.6 μg/ml) was not significantly different from that in control goats (55.4 μg/ml).     

Ultrafiltration-light absorption spectrophotometric determination of silicon in blood

An ultrafiltration-light absorption spectrometric method for soluble molybdate-reactive silicon was assessed and applied to bovine and ovine blood plasma and sera, giving precise analytical results. Interfering protein above molecular weight 10,000–25,000 was removed by ultrafiltration, and silicon in ultrafiltrates was quantitated by measuring light absorption at 810 nm of the 1,2,4-aminonaphthol sulfonic acid/ascorbic acid-reduced silicomolybdate. Chemical interferences on the color-forming reaction of remaining blood components were tested by measuring recoveries of silicon added to real blood plasma samples and to synthetic blood plasma solutions, the latter containing typical levels of the major ions Na⁺, K⁺, Ca²⁺, HCO₃^?, and Cl^?, together with varying quantities of the potential interferants (amount per analytical reaction): phosphate (0–0.5 mg P), ferric ion (0–3 mg), fluoride (0–1.25 mg), vanadate (0–0.5 mg V), arsenate (0–10 μg As), and germanate (0–0.5 μg Ge). The mean recovery of added 0.8–9 μg silicon/g of bovine and ovine plasma was 97.7% (SE = 1.0, n = 17); the mean recovery of 1 and 5 μg silicon from synthetic blood plasma solutions with interferant levels up to 50-fold that in normal plasma was 99.2% (SE = 0.3, n = 47). Silicon concentrations found in bovine and ovine blood plasma and sera were typically around 7 μg/ml with procedural reagent blanks consistently low at a mean of 0.12 μg/test (SD = 0.011, n = 20). The silicon level in Center for Disease Control bovine serum (reference specimen Lot R-2274) was found to be (mean ± SE, n = 10) 1.147 ± 0.013 μg/g or 1.172 ± 0.013 μg/ml (25°C). The method detectivity (detection limit) was estimated at 0.03 μg.     

Antibiofilm and antimicrobial activities of green synthesized silver nanoparticles using marine red algae Gelidium corneum

In our study, green synthesis of silver nanoparticles was carried out using a red algae Gelidium corneum extract as reducing agent. The obtained silver nanoparticles were characterized by UV–vis, TEM, XRD, FTIR and ICP-MS measurements. FTIR measurements indicated the possible functional groups responsible for the stabilization and reduction of nanoparticles, while XRD analysis results explained the crystalline structure of the particles with centric cubic geometry. TEM micrographs showed that the size of the nanoparticles was between 20–50 nm. According to the broth microdilution test results, AgNPs showed a high antimicrobial activity with very low MIC values (0.51 μg/ml for Candida albicans yeast and 0.26 μg/ml for Escherichia coli bacteria). The different ultrastructural effects of silver nanoparticles on yeast and bacterial cells were observed by TEM. Antibiofilm efficacy studies were also examined in two stages as prebiofilm and postbiofilm effect. In prebiofilm effect studies, AgNPs (0.51 μg/ ml) exhibited 81% reducing effect on biofilm formation. The highest reduction rate in postbiofilm studies was 73.5% and this was achieved with 2.04 μg/ml AgNPs. Our data support that the silver nanoparticles obtained by this environmentally friendly process have potential to be used for industrial and therapeutic purposes.     

The effect of purified alpha-fetoprotein on in vitro assays of cell-mediated immunity.

We have evaluated the effect of purified human α-fetoprotein (AFP) on several in vitro correlates of cell-mediated immunity. AFP (100 μg/ml) had no effect on antigen-induced migration inhibitory factor (MIF) production or on the ability of T cells to bind sheep erythrocytes. In contrast, AFP had a twofold effect on mitogen-and antigen-induced lymphocyte proliferation. In a dose-dependent fashion (1–100 μg/ml), AFP was mitogenic to lymphocyte cultures and also suppressed tritiated thymidine incorporation by PHA- and SK-SD-stimulated cultures. Albumin had somewhat similar effects on lymphocyte proliferation but only at 100 μg/ml. The reason for the latter was not clear since the albumin preparation was free of any detectable AFP. Our studies suggest that human AFP may not have any biologically significant immunosuppressive function.     

Cortisol and cortisone levels in umbilical cord plasma and maternal plasma of normal pregnancies

A method for assaying cortisol and cortisone using chromatography on either paper or Sephadex LH-20 columns for isolation, followed by competitive protein binding, has been applied to umbilical cord and maternal plasma samples. In mixed cord plasma the mean cortisol concentration was 6.0 ± 0.8 μg/100 ml (n = 9) and the mean cortisone concentration was 13.5 ± 2.9 μg/100 ml (n = 9). In cord arterial plasma the mean cortisol concentration was 6.3 ± 2.9 μg/100 ml (n = 6) and the mean cortisone level was 10.1 ± 2.5 μg/100 ml (n = 6). For cord venous plasma, the mean level of cortisol was 5.6 ± 1.5 μg/100 ml (n = 6) and of cortisone was 13.5 ± 2.4 μg/100 ml (n = 6). Maternal plasma gave a mean value of cortisol of 42.3 ± 4.5 μg/100 ml (n = 6) and of cortisone of 6.2 ± 0.9 μg/100 ml. The results of this study suggest that the fetus at term-gestation produces cortisol. The significance of this production compared with placental transfer of maternal cortisol into the fetal circulation however is uncertain.     

The effect of cholecystokinin octapeptide on pituitary-adrenal hormone secretion

The purpose of this investigation was to determine the influence of cholecystokinin octapeptide (CCK-OP) on pituitary-adrenal hormone secretion. CCK-OP at a dose of 5 μg/kg (i.p.) elevated plasma corticosterone from 27 to 43 μg/100 ml in one experiment and from 12 to 50 μg/100 ml in a second experiment: Lower doses of CCK-OP (0.5 μg/kg) elevated corticosterone from 12 μg/100 ml to 20 μg/100 ml. CCK-OP (1, 10, and 100 ng/ml) had no effect on ACTH-induced corticosterone released by isolated adrenal cells in vitro when tested in the presence of 50 pg of ACTH_1?24. 100 and 500 ng of CCK-OP resulted in an increased pituitary ACTH release equal to 123% (n.s.) and a 206% (P < 0.05) of control, respectively. In comparison, a $\text{3}\text{5}$ hypothalamic stalk median eminence equivalent increased ACTH release to 313% of control (P < 0.05). The exact mechanism of this CCK effect on pituitary ACTH release is unknown. Although it is likely that the direct effects on the pituitary in vitro represent a pharmacologic and not a physiologic effect of this peptide, in vivo doses are between doses used for pancreatic effects and satiety effects suggesting that there may be a physiologic stimulating action of this peptide on the hypothalamic-pituitary-adrenal axis but at a level above the adrenal and pituitary.     

Enantioselective determination of sotalol enantiomers in biological fluids using high-performance liquid chromatography

A simple and sensitive high-performance liquid chromatograhic (HPLC) method for the determination of (+)-(S)-sotalol and (−)-(R)-sotalol in biological fluids was established. Following extraction with isopropyl alcohol from biological samples on a Sep-Pak C₁₈ cartridge, the eluent was derivatized with 2,3,4,6-tetra-O-acetyl-β-d-glucopyranosol isothiocyanate (GITC). The diastereoisomeric derivatives are resolved by HPLC with UV detection at 225 nm. Calibration was linear from 0.022 to 4.41 μg/ml in human plasma and from 0.22 to 88.2 μg/ml in human urine for both (+)-(S)- and (−)-(R)-sotalol. The lower limit of determination was 0.022 μg/ml for plasma and 0.22 μg/ml for urine. The within-day and day-to-day coefficients of variation were less than 7.5% for each enantiomer at 0.09 and 1.8 μg/ml in plasma and at 0.44 and 4.4 μg/ml in urine. The method is also applicable to other biological specimens such as rat, mouse and rabbit plasma.     

Effect of tumor promoter 12-O-tetradecanoyl-phorbol 13-acetate on induction of sister-chromatid exchanges in Chinese hamster V79 cells treated with mutagens

The effect of a tumor promoter, 12-O-tetradecanoyl-phorbol 13-acetate (TPA) alone and in combination with mitomycin C (MMC) or cyclophosphamide (CPP) on the induction of sister-chromatid exchanges (SCE) in Chinese hamster V79 cells was investigated. TPA alone at various doses and durations caused no increase of SCE frequency. MMC either at the dose of 0.03 or 0.003 μg/ml alone or in combination with TPA (2 μg/ml) all caused a significant increase of SCE frequencies. There was no difference in SCE frequencies between the cultures treated with MMC alone at 0.03 μg/ml and those treated with MMC plus TPA. However, cultures treated with MMC at 0.003 μg/ml plus TPA had significantly and consistently higher SCE frequencies than those treated with MMC alone at all durations. Treatment of CPP at 1 μg/ml activated by S9 mix caused significant increase of SCE frequencies. Surprisingly, the cultures treated with CPP, S9 mix plus TPA (2 μg/ml) caused a drastic reduction of SCE frequencies as compared to those treated with CPP and S9 mix only at all durations. These results indicate that TPA alone had no effect on SCE in V79 cells. TPA enhanced the SCE induction in V79 cells treated with MMC at a low dose, i.e. 0.003 μg/ml, but it inhibited SCE induction in cultures treated with the indirect mutagen CPP. Thus, TPA has no direct effect on genetic materials but it may indirectly alter the effects of a mutagen.